G protein-coupled receptors detect many different external signals including ones that regulate gene expression and cellular differentiation. While many components of G protein-mediated signal transduction pathways are known, the mechanisms by which different signals regulate specific genes remains to be determined. Pathway specificity is in part determined by the G protein G(alpha) subunits and recent data indicate these subunits play a role in gene regulation in the soil amoebae Dictyostelium. The goal of the proposed research is to test the hypothesis that G(alpha) subunits regulate gene expression through interactions with other signaling components, such as MAP (mitogen-activated protein) kinases. To achieve this goal, the early developmental expression of discoidin (Dsc) in Dictyostelium will be examined in cells lacking the MAP kinase Erk2 and expressing different wild-type or chimeric G(alpha) subunits. In addition, a genetic screen will be used to identify suppressor mutations that rescue pDSC/GFP reporter gene expression from the repression of a chimeric G(alpha) subunit. The suppressor mutations will be created by REMI mutagenesis, allowing genes to be tagged that might function downstream of G(alpha) subunits to regulate gene expression. Additional characterization of suppressor genes will be conducted to verify their function regulating DSC gene expression downstream of G(alpha) subunits in these signaling pathways. These studies will provide significant insights into the signaling mechanisms that control the differentiation of eukaryotic cells.